Generally, when researching tiny regions of interest on a sample, researchers often employ a microscope to observe the sample. The microscope may be a conventional wide-field, fluorescence, epi-fluorescence or confocal microscope. The optical configuration of such a microscope typically includes a light source, illumination optics, objective lens, sample holder, imaging optics and a detector. Light emitted from the light source illuminates the region of interest on the sample after propagating through the illumination optics and the objective lens. Microscope objective forms a magnified image of the object that can be observed via eyepiece, or in case of a digital microscope, the magnified image is captured by the detector and sent to a computer for live observation, data storage, and further analysis.
The light source sends an excitation light that reaches the objective of an epi-fluorescence microscope that should ideally completely fill-in its back aperture. However, due to the design of the objective lenses these lenses have different numerical aperture (NA) that have different aperture diameters, which under conditions of a constant beam diameter may prevent the back aperture from being completely filled. Under-filling of the objective aperture leads to a smaller effective NA and thereby degradation of the lateral and axial resolution in general, and of optical sectioning performance for confocal systems. Over-filling of the objective aperture only uses a fraction of the excitation energy and results in decreased illumination efficiency.
There have been prior patents, such as U.S. Pat. No. 6,081,371 where for point-confocal and widefield microscopes the excitation beam diameter can be easily adjusted to match different objectives using a conventional zoom system. For these patents, the beam is circularly symmetric about the optical axis and the zooming optical elements maintain that symmetry. In contrast, in a line-confocal system the beam lacks circular symmetry and a specialized zooming solution is required. The application of a point confocal zooming system, as shown in this patent, to a line-confocal microscope is undesired because it leads to significant image non-uniformity.
With regard to other prior patents, such as U.S. Pat. No. 7,235,777 a complex design is proposed to achieve near one-dimensional zooming. However, this patent is a complex design that does not provide an optimized illumination system for all of the supported objectives. Therefore, there is a need for a simple adjustable system that provides optimized illuminations of back apertures for all the supported objectives.